Direct measurement of RNA Polymerase II hypertranscription in cancer FFPE samples

Hypertranscription occurs at targets of oncogene transcription factors in aggressive human cancers. However, detection relies on mRNAs which are heavily processed and have variable half-lives, and on accurate cell number estimations. Here we apply FFPE-CUTAC on slides and curls to quantify hypertranscription at regulatory elements and replication-coupled histone genes. Overall design: We used FFPE-CUTAC, a modification of the CUT&Tag chromatin profiling strategy, whereby antibody-targeted controlled integration of DNA sequencing adapters at sites of paused RNA Polymerase II or between nucleosomes flanking open chromatin sites produces libraries for paired-end sequencing from Formalin-Fixed Paraffin-Embedded (FFPE) sections.

过度转录（Hypertranscription）发生于侵袭性人类癌症中致癌基因转录因子的靶标区域。然而，当前的检测手段依赖于经过高度加工、半衰期存在差异的信使RNA（messenger RNA, mRNA），同时还需进行精确的细胞数量估算。本研究将FFPE-CUTAC技术应用于组织切片与石蜡卷样本，以定量分析调控元件及复制偶联组蛋白基因区域的过度转录水平。实验整体设计：本研究采用FFPE-CUTAC技术——这是对CUT&Tag染色质图谱分析策略的改良方案——其核心原理为，在暂停状态的RNA聚合酶II（RNA Polymerase II）结合位点，或是开放染色质区域侧翼的核小体之间，通过抗体靶向实现DNA测序接头的可控整合，进而从福尔马林固定石蜡包埋（FFPE）组织切片中构建出可用于双端测序的文库。